Mammalian and Drosophila PCNA accumulate at RF interchangeably between these divergent organisms. Drosophila S2 and mammalian C2C12 cells were transfected, pulse labeled with BrdU to label sites of active DNA replication, and fixed with formaldehyde followed by immunostaining. (A) S2 cells expressing HsPCNA and coimmunostained with anti-PCNA (FL-261; Santa Cruz) and anti-BrdU antibodies. Anti-PCNA antibody (FL-261) specifically stains transfected cell (arrow) expressing HsPCNA. (B) S2 cells expressing HsPCNA (top) or GFP-HsPCNA (bottom) and immunostained with anti-BrdU. HsPCNA was detected as in (A). Overlay shows colocalization of HsPCNA and GFP-HsPCNA with BrdU foci. (C) C2C12 cells transfected with plasmid encoding GFP-DmPCNA and immunostained with anti-BrdU. Scale bar = 2 μm.